Abstract
Cdc7 is a serine-threonine kinase that regulates initiation and progression of DNA replication. The activity of purified Cdc7 kinase is significantly stimulated by polyamines such as spermine or spermidine. Positively charged polymers of lysine or arginine also stimulate its kinase activity, whereas the negatively charged substances such as polyglutamate or nucleic acids significantly inhibit the kinase activity. Spermine affects both the K(m) and V(max) of Cdc7 kinase for a minichromosome maintenance (MCM) substrate. We also found that histones, lysine- and arginine-rich basic proteins, can stimulate Cdc7 kinase activity, and a MCM complex in association with histone is a more efficient substrate of Cdc7 than the free MCM complex. These results identify potential cellular inhibitors and stimulators of Cdc7 kinase and suggest that Cdc7 may be another target of cellular polyamines and that histones may stimulate Cdc7-mediated phosphorylation of chromatin-bound substrates. Ectopic expression of an antizyme, known to reduce the cellular polyamine levels, resulted in reduction of Cdc7-mediated phosphorylation of MCM4 protein, suggesting physiological roles of polyamines in regulation of Cdc7 kinase activity in the cells.
Highlights
Cdc7 kinase is composed of two subunits, namely Cdc7-related catalytic subunit (Hsk1 in fission yeast) and Dbf4-related activation subunit
The results indicate that Cdc7 kinase is stimulated by positively charged polymers such as spermine or polylysine or histones and is inhibited by negatively charged polymers including nucleic acids
Cdc7 Kinase Activity Is Stimulated by Polyamines in Vitro— Previous results indicate that Cdc7 kinase may target serine/ threonine residues with an acidic environment [8, 9]
Summary
Cdc7 kinase is composed of two subunits, namely Cdc7-related catalytic subunit (Hsk1 in fission yeast) and Dbf4-related activation subunit The results indicate that Cdc7 kinase is stimulated by positively charged polymers such as spermine or polylysine or histones and is inhibited by negatively charged polymers including nucleic acids. The substrates were mouse MCM2-4-6-7 (0.8 g) or Mrc1 protein (0.1 g) for human Cdc7-ASK and for fission yeast Hsk1-Dfp1/Him1 complex.
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